Chemical characterization of the regularly arranged surface layers of Clostridium thermosaccharolyticum and Clostridium thermohydrosulfuricum

Abstract

Clostridum thermosaccharolyticum and Clostridiurm thermohydrosulfuricum possess as outermost cell wall layer a tetragonal or hexagonal ordered array of macromolecules. The subunits of the surface layer can be detaqhed from isolated. cell walls with urea (8 M) or guanidine-HCl (4 to 5 M). Triton X-100, dithiothreitol, ethylenediaminetetraacetate, and KCl (3 M) had no visible effect on the regular arrays. Sodium dodecyl sulfate-polyacrylamide electrophroesis showed that, in both organisms, the surface layer is composed of glycoprotein of molecular weight 140,000. The glycoprotein from both microorganisms has a predominantly acidic amino acid composition and an acidic isoelectric point after isoelectric focusing on polyacrylamide gels. The glycocomponent is composed of glucose, galactose, mannose, and rhamnose.

Chemical characterization of the regularly arranged surface layer glycoprotein of Clostridium thermosaccharolyticum D120-70

“…Freeze-etched preparations of intact cells of C. thermosaccharolyticum showed the presence of a square surface-layer lattice ( Fig. 1 a), as already demonstrated by Sleytr and Thorne [5]. Thin sections confirmed the location of the surface layer as the outermost cell envelope component ( Fig.…”

“…Growth of bacteria and isolation of the glycopeptide C. thermosaccharolyticum D120-70 was grown as described and the surface-layer glycoprotein was isolated from clean cell walls by extraction with 5 M guanidine hydrochloride [5]. Upon thorough dialysis of this extract three times against 2 1 distilled water, self-assembly products consisting of pure surface-layer glycoprotein subunits were formed [5]. The glycopeptides were obtained after pronase digestion (1 5% pronase) as described previously [8].…”

“…and --+ 4)-fl-~-GlcpNAc-(l+ 3)-P-~-ManpNAc-(l-+ t Regularly arranged surface layers have been found in nearly every taxonomic group of walled eubacteria and represent an almost universal feature of archaebacterial cell envelopes [l, 21. The chemical composition of the surface layer glycoprotein of the archaebacterium Halobacterium halobium has been described by Wieland and coworkers in great detailIn the course of the structural characterization of the surface layers of the taxonomically closely related thermophilic eubacteria Clostridium thermosaccharolyticum D120-70 and Clostridium thermohydrosulfuricum Ll l l-69 [4], chemical analysis provided first evidence for the presence of glycoprotein subunits in the surface layer arrays of these strains [5]. Since then the structures of the carbohydrate portions of surface-layer glycoproteins of C. thermohydrosulfuricum Llll-69 [6] and Bacillus stearothermophilus NRS 2004/3a [7, 81 have been elucidated and the nature of the linkage region of one of these glycopeptides has been described [9].…”

“…Oneand two-dimensional nuclear magnetic resonance studies, together with methylation analysis and periodate oxidation, were used to determine the structures of the polysaccharide portions of these glycopeptides. The combined chemical and spectroscopic evidence suggests the following structures : In the course of the structural characterization of the surface layers of the taxonomically closely related thermophilic eubacteria Clostridium thermosaccharolyticum D120-70 and Clostridium thermohydrosulfuricum Ll l l-69 [4], chemical analysis provided first evidence for the presence of glycoprotein subunits in the surface layer arrays of these strains [5]. Since then the structures of the carbohydrate portions of surface-layer glycoproteins of C. thermohydrosulfuricum Llll-69 [6] and Bacillus stearothermophilus NRS 2004/3a [7, 81 have been elucidated and the nature of the linkage region of one of these glycopeptides has been described [9].…”

A high‐molecular‐mass cell wall protein released from Clostridium tyrobutyricum by heat treatment

“…The localization of the high-molecular-mass component of the cell wall of C. tyrobutyricum is not yet known. However, many bacteria including Clostridium species are known to possess regular arrays composed of hexagonally or tetragonally arranged protein subunits of high molecular mass on the outer surface of their cell walls [15], Authors in [16] observed an ordered layer of glycoprotein of 140 kDa in C. tbermosaccbaro~yticum and thermohydrosuffuricum and those in f 171 also observed an ordered layer of protein of 195 kDa on C.botulinum type A. In the case of C. tyrobutyricum, such regular arrays have not yet been demonstrated but the high-molecular-mass component found in its cell wall may perhaps belong to a structure of this type.…”